TRANSGENESIS

=Transgenesis= ''(also called genetic modification)'' [image:http://i.imgur.com/J6peQHk.png] Transgenesis is the process of introducing a gene (referred to as a transgene) from one organism into the genome of another organism. The aim is that the resulting transgenic organism will express the gene and exhibit some new property or characteristic. This is made possible by the fact that the genetic code is universal for all living things. Steps involved in the process of transgenesis are outlined below. ===1. IDENTIFICATION=== A gene that codes for a desirable trait or protein must first be identified . Researchers may identify desirable traits in other species and try to identify the gene responsible. There are several techniques that can be used to identify the gene sequence that codes for the specific protein / trait if interest. If the protein has been isolated it may be possible to determine its amino acid sequence (or part there of). If the amino acid sequence is know it may be possible to determine part of the gene sequence using a codon table, however there will usually be several different possible sequences due to the redundancy of the genetic code. Once a small part of the sequence has been determined it may be possible to construct a [https://www.pathwayz.org/Tree/Plain/1004 DNA probe] that will stick the target gene. Other techniques that may be used to identify a gene include: '''Gene Chips (Microarrays)''' and '''DNA Sequencing''' ===2. ISOLATION=== The target gene must then be isolated. DNA from the organism that contains the target gene can usually be isolated simply by breaking up cells mechanically or with chemical treatments such as detergents. The DNA can be separated from the other cell components using a technique call '''centrifugation'''. To separate the target gene from the rest of the DNA it would first be cut using a [https://www.pathwayz.org/Tree/Plain/1001 restriction enzyme]. The fragments would then be separated according to size using a technique called [https://www.pathwayz.org/Tree/Plain/1008 Gel Electrophoresis] The fragment tat contains the target gene can be identified using a [https://www.pathwayz.org/Tree/Plain/1004 DNA probe] And can then cut out of the gel and amplified (copied) using [https://www.pathwayz.org/Tree/Plain/1003 PCR] Alternatively the gene could be inserted into a bacterial plasmid using [https://www.pathwayz.org/Tree/Plain/1109 DNA Ligase] The bacteria would then copy the gene each time underwent cell division (a technique called [https://www.pathwayz.org/Tree/Plain/1005 Gene Cloning]) If enough is know about the DNA sequence either side of the gene it may be possible to make specific DNA primers and copy the gene using PCR without first isolating it on a gel. ===3. TRANSFORMATION=== A vector is then used to transfer the target gene (transgene) into the organism being modified. There are many different vectors / techniques used to transfer the transgene depending on the cell type etc. See [https://www.pathwayz.org/Tree/Plain/1007 Transgenesis - Vectors] for some examples. However, if the gene is inserted on it's own it is unlikely that it will be expressed. Special ''promotor'' and ''termination'' sequences might be needed either side of the gene in order for it to be expressed. The type of promoter might determine where (in which tissues) the protein is expressed. The final DNA sequence that is prepared including the target gene and associated regulatory sequences is called a ''Gene Construct'' Most vectors / techniques have low success rates, only producing a very small percentage of cells that actually express the transgene. In order for the gene to be expressed it must make it's way into the nucleus. For it to be passed on during cell division (mitosis and meiosis) it must integrate into the target cells genome (usually by recombination - crossing over). For this reason scientists often incorporate a '''reporter gene''' into the gene construct. This is a second gene that codes for an easily selectable / observable characteristic e.g. antibiotic resistance or glow in the dark protein. This makes it easier for researchers to establish while cells have successfully integrated and are expressing the transgene. ==Case Study: Insulin [extra for experts]== Insulin is a protein hormone that is needed to regulate blood sugar levels. Many diabetics do not produce adequate levels of insulin own their own. Originally insulin was isolated from pigs; however, this had major draw-backs (cost, risk of infection, immune reaction). Once the gene for human insulin had been isolated, it was cloned using bacteria (see gene cloning). However, the bacteria did not simply copy the gene, they also expressed it. The bacteria were cultured under optimum growth conditions and the insulin was extracted. [image:http://i.imgur.com/okNJHAN.png] In reality it wasn't quite so simple. Gene expression is controlled by regulatory regions such as promoters, etc (promoters control whether a gene is switched on or off). Promoters that would increase the expression of the insulin gene also needed to be incorporated. Also, Insulin is actually made of two protein chains that are held together by disulphide bridges (a type of chemical bond). There are actually two genes that each code for one of the two protein chains. Thus the whole process had to be done twice, once for each gene. The disulphide bridges that hold the two strands together could then, quite simply, be incorporated chemically. This is an example of a post-translational modification - a change that is made after the protein chain(s) have been synthesized. While this modification could quite easily be replicated chemically in the lab, it is not always quite so simple. Many eukaryote proteins undergo more complex post translational modifications such as methylation (have methyl groups added to specific amino acids). These cannot be done in the lab, nor are bacteria capable performing these modifications to the protein chains (as they lack the necessary organelles –endoplasmic reticulum, golgi apparatus, etc). In order to carry out the post translational modifications, the proteins must be made within eukaryote cells, which have the necessary machinery / organelles to carry out these modifications. ==Implications of Transgenesis== Transgenesis can have implications on: • '''Genetic Biodiversity''' • '''Ecosystems''' • '''The health or survival of individuals''' • '''The survival of populations''' • '''The evolution of populations''' E.g. Transgenesis may be used to generate a need breed of animals that is genetically different thus contributing to genetic diversity. In some cases the transgenic protein may have side effects on the genetically modified animal. This could have a positive or negative impact on the animals health. The implications are specific to each each and depend on the gene being added (or removed), the organism being modified and it's environment.